scholarly journals A simple graphical method for determining the inhibition constants of mixed, uncompetitive and non-competitive inhibitors (Short Communication)

1974 ◽  
Vol 137 (1) ◽  
pp. 143-144 ◽  
Author(s):  
Athel Cornish-Bowden
Keyword(s):  
Short Communication ◽  
Graphical Method ◽  
Competitive Inhibitor ◽  
Inhibition Constant ◽  
New Method ◽  
Competitive Inhibitors ◽  
Inhibition Constants ◽  
Inhibited Enzyme

A new method is described for plotting kinetic results for inhibited enzyme-catalysed reactions. It provides a simple way of determining the inhibition constant, K′i, of an uncompetitive, mixed or non-competitive inhibitor.

Di- and tripeptide N-alkyl- and N-aralkyl amides as chymotrypsin inhibitors

1988 ◽  
Vol 53 (11) ◽  
pp. 2877-2883 ◽  
Author(s):  
Evžen Kasafírek ◽  
Irena Šutiaková ◽  
Michal Bartík ◽  
Antonín Šturc
Keyword(s):  
Methyl Ethyl ◽  
Competitive Inhibitors ◽  
Inhibition Constants ◽  
Chymotrypsin Inhibitors

Two competitive inhibitors of chymotrypsin, Glt-Ala-Ala-Leu-EtPh and Glt-Ala-Ala-Pro-NH-EtPh, were synthesized and their inhibition constants Ki were determined. The Ki-determination was carried also with a set of peptides of type X-(Ala)nNH-Y, where X is 3-carboxypropionyl- or 4-carboxybutyryl-, n is 2 or 3 and Y is methyl, ethyl, diethyl, isopropyl, propyl, butyl, isobutyl and 2-phenylethyl. Chymotrypsin inhibition was observed only with peptides containing an aralkyl residue whereas peptides with an alkyl are without any effect. Glt-Ala-Ala-Leu-NH-EtPh shows the highest Ki-value (80 μmol l-1).

Receptor kinetics pertaining to blockade of nerve-released transmitter at synapses

1988 ◽  
Vol 233 (1273) ◽  
pp. 461-475 ◽  
Keyword(s):  
Channel Blocker ◽  
Competitive Inhibitor ◽  
Tissue Response ◽  
Receptor Density ◽  
Receptor Kinetics ◽  
Tissue Responses ◽  
Competitive Inhibitors ◽  
Transmitter Substance ◽  
Uncompetitive Inhibitor ◽  
Temporal Inhomogeneity

The question is raised as to whether competitive inhibitors should block responses of tissue to nerve-released neurotransmitter to the same extent as they block equivalent responses to exogenous agonist. From a simple dynamic model of synaptic events, which takes into account non-constancy of transmitter concentration in space and time, it is deduced that equal blockade of responses to nerve-released and exogenous transmitter substance will occur if: (i) there are locally many more receptor molecules than transmitter molecules; (ii) the active agonist–receptor complex, A n R, has n = 1 ; and (iii) tissue response is insensitive to spatial or temporal inhomogeneity of AR. In such a case there will also be equal sensitivity of responses to other modes of inhibition: irreversible competitive, uncompetitive, and non-competitive. Equal blockade of responses to equi-effective endogenous and exogenous agonist will also occur if nerve stimulation gives rise to a steady uniform concentration of agonist, so that equilibrium kinetics are applicable. When n > 1 and/or when tissue responses reflect local peak A n R, response to nerve-released transmitter will be relatively insensitive to receptor blockade by a competitive inhibitor. The same is true for irreversible competitive blockade or for modulation of receptor density. However, an uncompetitive inhibitor (e. g. a ‘channel blocker’) may be more effective against nerve-released agonist than against exogenous agonist.

A New Method for the Construction of Hencky-Prandtl Nets

1957 ◽  
Vol 24 (1) ◽  
pp. 81-84
Author(s):  
E. G. Thomsen
Keyword(s):  
Graphical Method ◽  
New Method ◽  
Physical Plane ◽  
Rapid Construction

Abstract A graphical method for the rapid construction of Hencky-Prandtl nets has been developed. The principle employed has been stated in a theorem. It is shown that the method may be used for nets on both the physical plane and on the velocity plane or hodograph. The use is illustrated by examples.

Recognition of Short Time Periods through Seriation

1977 ◽  
Vol 42 (4) ◽  
pp. 628-629 ◽  
Author(s):  
Clement W. Meighan
Keyword(s):  
Recent Article ◽  
Graphical Method ◽  
New Method ◽  
Time Discrimination ◽  
Time Periods ◽  
Short Time

One aspect of the recent article by Drennan (1976) merits some additional discussion. This has to do with the units of time that can be discriminated by seriation methods. One advance claimed for the new method proposed is that it allows for time placement within 25 years or so, stated as “ … finer than most traditional seriation.” In an article published 17 years ago, I showed this degree of time discrimination, with a much simpler graphical method of seriation.

scholarly journals Mechanism of d-fructose isomerization by Arthrobacterd-xylose isomerase

1992 ◽  
Vol 283 (1) ◽  
pp. 223-233 ◽  
Author(s):  
M Rangarajan ◽  
B S Hartley
Keyword(s):  
Ring Opening ◽  
Ph Dependence ◽  
Xylose Isomerase ◽  
Competitive Inhibitor ◽  
Apparent Dissociation Constant ◽  
Ph Values ◽  
Rate Limiting Step ◽  
Competitive Inhibitors ◽  
Rate Limiting ◽  
Very High

The mechanism of D-fructose isomerization by Arthrobacter D-xylose isomerase suggested from X-ray-crystallographic studies was tested by detailed kinetic analysis of the enzyme with various metal ions at different pH values and temperatures. At D-fructose concentrations used in commercial processes Mg2+ is the best activator with an apparent dissociation constant of 63 microM; Co2+ and Mn2+ bind more strongly (apparent Kd 20 microM and 10 microM respectively) but give less activity (45% and 8% respectively). Ca2+ is a strict competitive inhibitor versus Mg2+ (Ki 3 microM) or Co2+ (Ki 105 microM). The kinetics show a compulsory order of binding; Co2+ binds first to Site 2 and then to Site 1; then D-fructose binds at Site 1. At normal concentrations Mg2+ binds at Site 1, then D-fructose and then Mg2+ at Site 2. At very high Mg2+ concentrations (greater than 10 mM) the order is Mg2+ at Site 1, Mg2+ at Site 2, then D-fructose. The turnover rate (kcat.) is controlled by ionization of a residue with apparent pKa at 30 degrees C of 6.0 +/- 0.07 (Mg2+) or 5.3 +/- 0.08 (Co2+) and delta H = 23.5 kJ/mol. This appears to be His-219, which is co-ordinated to M[2]; protonation destroys isomerization by displacing M[2]; Co2+ binds more strongly at Site 2 than Mg2+, so competes more strongly against H+. The inhibition constant (Ki) for the two competitive inhibitors 5-thio-alpha-D-glucopyranose and D-sorbitol is invariant with pH, but Km(app.) in the Mg[1]-enzyme is controlled by ionization of a group with pKa 6.8 +/- 0.07 and delta H = 27 kJ/mol, which appears to be His-53. This shows that Km(app.) is a complex constant that includes the rate of the ring-opening step catalysed by His-53, which explains the pH-dependence. In the Mg[1]Mg[2]-enzyme or Co[1]Co[2]-enzyme, the pKa is lower (6.2 +/- 0.1 or 5.6 +/- 0.08) because of the extra adjacent cation. Hence the results fit the previously proposed pathway, but show that the mechanisms differ for Mg2+ and Co2+ and that the rate-limiting step is isomerization and not ring-opening as previously postulated.

The copurification of β-glucosidase, β-xylosidase, and 1,3-β-glucanase in two separate enzyme complexes isolated from Trichoderma harzianum E58

1987 ◽  
Vol 65 (9) ◽  
pp. 822-832 ◽  
Author(s):  
Larry U. L. Tan ◽  
Paul Mayers ◽  
Michelle Illing ◽  
John N. Saddler
Keyword(s):  
Isoelectric Point ◽  
Mercuric Chloride ◽  
Trichoderma Harzianum ◽  
Inhibition Constant ◽  
Activity Ratios ◽  
Molecular Masses ◽  
Inhibition Constants ◽  
Inhibition Studies ◽  
Enzyme Complexes

Two enzyme complexes, each with β-glucosidase (β-D-glucoside glucohydrolase, EC 3.2.1.21), β-xylosidase (β-D-xylan xylohydrolase, EC 3.2.1.37), and 1,3-β-glucanase (laminarinase, EC 3.2.1.39) activity, were purified to near homogeneity from the cellulolytic fungus Trichoderma harzianum E58. The two complexes had the same isoelectric point of pH 8.3 and identical subunit molecular masses of 75 400 daltons. The two complexes were also similar in that all activities were sensitive to inhibition by mercuric chloride (2 mM) and D-glucono-1,5-lactone (0.2% w/v). The activity ratios of the major and minor complexes were 1:1.7:4.3 and 1:1.6:3.1 for the β-xylosidase, β-glucosidase, and 1,3-β-glucanase, respectively. Both complexes had approximately the same Km values for p-nitrophenyl β-D-glucopyranoside and salicin. The pH optima of corresponding activities of the two complexes were also similar. The major and minor complexes differed in that the Km of the former for laminarin was almost threefold lower than that of the latter. Whereas all three activities of the minor complexes were inhibited by D-glucono-1,5-lactone with the same inhibition constant, the β-glucosidase and 1,3-β-glucanase of the major complex had inhibition constants which differed by more than 80 000 times. In addition, the inhibition on the 1,3-β-glucanase in the major and minor complexes using D-glucono-1,5-lactone were noncompetitive and competitive, respectively. From the inhibition studies, the β-glucosidase, β-xylosidase, and 1,3-β-glucanase activities in the minor complex were deduced to be more interdependent than the same activities in the major complex.

scholarly journals Structural modeling of GSK3β implicates the inactive (DFG-out) conformation as the target bound by TDZD analogs

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Meenakshisundaram Balasubramaniam ◽  
Nirjal Mainali ◽  
Suresh Kuarm Bowroju ◽  
Paavan Atluri ◽  
Narsimha Reddy Penthala ◽  
...  
Keyword(s):  
Glycogen Synthase ◽  
Three Dimensional ◽  
Binding Mode ◽  
Competitive Inhibitor ◽  
Dimensional Structure ◽  
High Potency ◽  
Subsequent Formation ◽  
Inactive Conformation ◽  
Competitive Inhibitors ◽  
Physiological Pathways

Abstract Glycogen synthase kinase-3β (GSK3β) controls many physiological pathways, and is implicated in many diseases including Alzheimer’s and several cancers. GSK3β-mediated phosphorylation of target residues in microtubule-associated protein tau (MAPTAU) contributes to MAPTAU hyperphosphorylation and subsequent formation of neurofibrillary tangles. Inhibitors of GSK3β protect against Alzheimer’s disease and are therapeutic for several cancers. A thiadiazolidinone drug, TDZD-8, is a non-ATP-competitive inhibitor targeting GSK3β with demonstrated efficacy against multiple diseases. However, no experimental data or models define the binding mode of TDZD-8 with GSK3β, which chiefly reflects our lack of an established inactive conformation for this protein. Here, we used metadynamic simulation to predict the three-dimensional structure of the inactive conformation of GSK3β. Our model predicts that phosphorylation of GSK3β Serine9 would hasten the DFG-flip to an inactive state. Molecular docking and simulation predict the TDZD-8 binding conformation of GSK3β to be inactive, and are consistent with biochemical evidence for the TDZD-8–interacting residues of GSK3β. We also identified the pharmacophore and assessed binding efficacy of second-generation TDZD analogs (TDZD-10 and Tideglusib) that bind GSK3β as non-ATP-competitive inhibitors. Based on these results, the predicted inactive conformation of GSK3β can facilitate the identification of novel GSK3β inhibitors of high potency and specificity.

Rat liver L-threonine deaminase: Properties and purification

1985 ◽  
Vol 5 (6) ◽  
pp. 499-508 ◽  
Author(s):  
R. Leoncini ◽  
R. Pagani ◽  
A. Casella ◽  
E. Marinello
Keyword(s):  
Thermal Stability ◽  
Rat Liver ◽  
Competitive Inhibitor ◽  
New Method ◽  
Optimum Temperature ◽  
Temperature Optimum ◽  
Threonine Deaminase ◽  
Ph Optimum ◽  
Carbonate Ions

A new method of purification of rat liver L-threonine deaminase has been developed, and the results obtained are compared with values obtained by other authors. Some properties of this enzyme (pH optimum, temperature optimum, thermal stability, specificity, etc.) have been examined and we found that the enzyme is inhibited by carbonate ions, that L-cysteine (a competitive inhibitor) is also an inactivator of the enzyme and that it is bound to the enzyme in a ratio of 0.25 mole of cysteine per mole of enzyme, supporting the hypothesis that the enzyme consists of 4 subunits.

scholarly journals The behaviour of trypsin towards α-N-methyl-α-N-toluene-p-sulphonyl-l-lysine β-naphthyl ester. A new method for determining the absolute molarity of solutions of trypsin

1968 ◽  
Vol 107 (1) ◽  
pp. 97-102 ◽  
Author(s):  
D. T. Elmore ◽  
J. J. Smyth
Keyword(s):  
Methyl Ester ◽  
Convenient Method ◽  
Substrate Concentration ◽  
Competitive Inhibitor ◽  
New Method ◽  
Spectrophotometric Measurement ◽  
Bovine Thrombin ◽  
The Absolute ◽  
Hydrolysis Of

1. α-N-Methyl-α-N-toluene-p-sulphonyl-l-lysine β-naphthyl ester (MTLNE) was synthesized as its hydrobromide and shown to be slowly hydrolysed by bovine pancreatic trypsin. The acylation step, however, is so much faster than deacylation of the acyl-enzyme that spectrophotometric measurement of the ‘burst’ of β-naphthol provides a convenient method for determining the absolute molarity of trypsin solutions. 2. By using the same stock solution of trypsin, application of this method at pH4·0 and pH7·0 as well as that of Bender et al. (1966) at pH3·7 gave concordant results. 3. Provided that [S]0>[E]0, the size of the ‘burst’ is independent of substrate concentration. 4. In the trypsin-catalysed hydrolysis of α-N-toluene-p-sulphonyl-l-arginine methyl ester, MTLNE functions as a powerful non-competitive inhibitor. 5. There is no detectable reaction between MTLNE and either bovine pancreatic α-chymotrypsin at pH4·0 or bovine thrombin at pH6·0.

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